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Abstract: FR-OR042

Comparative Analysis of Kidney Organoid and Adult Human Kidney by Single Cell RNA-Seq

Session Information

Category: Development, Stem Cells, and Regenerative Medicine

  • 501 Development, Stem Cells, and Regenerative Medicine: Basic

Authors

  • Wu, Haojia, Renal Division Washington University School of Medicine, Saint Louis, Missouri, United States
  • Uchimura, Kohei, Renal Division Washington University School of Medicine, Saint Louis, Missouri, United States
  • Donnelly, Erinn L., Renal Division Washington University School of Medicine, Saint Louis, Missouri, United States
  • Kirita, Yuhei, Renal Division Washington University School of Medicine, Saint Louis, Missouri, United States
  • Humphreys, Benjamin D., Renal Division Washington University School of Medicine, Saint Louis, Missouri, United States

Group or Team Name

  • The Humphreys Lab
Background

Kidney organoids differentiated from human pluripotent stem cells hold tremendous promise but the similarities and differences between protocols and between pluripotent cell sources is unknown. In addition, how closely organoid-derived kidney cell types model their adult human counterparts is undefined.

Methods

We used Dropseq to generate single cell transcriptomes from two different directed differentiation protocols (Morizane and Takasato), using both iPSC and hES cell lines for each. We analyzed and compared single cell gene expression from 71,390 cells isolated from 38 organoids using unbiased computational approaches. We could detect on average 1,115 genes per cell. We further compared the differentiation state of major organoid kidney cell types against 4,524 single nucleus transcriptomes that we generated from healthy adult human kidney.

Results

Both protocols generate kidney organoids that contain a diverse range of kidney cells at differing ratios but also substantial numbers of non-renal cell types such as muscle and neuron. Some organoids have up to 30% off-target cell types. Organoid-derived cell types are substantially immature compared with adult kidney cells. We could detect 16 different types of adult kidney cell types by single nucleus RNA-seq, including all glomerular cell types, all epithelial cell types including S1-S3 proximal tubule segments and type A and type B intercalated cells, as well as stroma and leukocytes. Disease-related genes predicted by GWAS are predominantly expressed in single cell types in adult kidney and organoids. We also identified lineage-specific expression of transcription factors, receptors and ligands during organoid differentiation and in adult human kidney.

Conclusion

We provide the first direct and comprehensive comparison of separate differentiation protocols using both iPSC and hES cells, revealing important differences in cell ratio, differentiation state and off-target cell types that will be important for investigators in the organoid field to appreciate. The information provided in this comprehensive dataset will also guide future attempts to improve differentiation protocols, including reduction of off-target populations.

Funding

  • NIDDK Support